The overall aim of this project is to elucidate the mechanisms that regulate the expression of the genes coding for proteins of the contractile apparatus in the human myocardium. To accomplish this goal we have selected the light chains of myosin for our initial studies. Myosin is the major myofibrillar protein, plays an important role in muscle contraction and exists in multiple molecular forms in mammalian cardiac and skeletal muscles. The expression of various isoforms of myosin shows a developmental and tissue-specific regulation, and for cardiac myosins, in particular, is modulated by various extrinsic factors. Preliminary studies indicate that the human ventricular light chains are indistinguishable from slow skeletal myosin light chains by several criteria. The ventricular light chains are also present in variable amounts in atria in normal human heart and this becomes more pronounced in cardiomyopathic hearts. Thus, the light chains are good candidates for studying the regulation of myofibrillar gene expression in the human myocardium. In this proposal we plan to study the structure and organization of the human ventricular, atrial and slow skeletal light chain genes. The cDNA and genomic probes will be used to examine the interrelationship of the genes and their tissue-specific expression. The 5' flanking sequences of the genes which are essential for transcriptional initiation and regulated expression will be analyzed. The role of various biochemical parameters such as genomic DNA methylation, chromatin sensitivity to nuclease, and interaction of effector molecules with 5' regulatory sequences, which are believed to be involved in eukaryotic gene regulation, will be examined in order to define the contribution of these parameters in the regulated expression of human cardiac light chain genes. It is hoped that the information obtained from these studies will provide a better understanding of the mechanisms of myofibrillar protein gene regulation in the normal and altered states of the human myocardium.